C1 inhibitor, a glycoprotein found in normal human serum, is a modulator of the activation of C1, the first component of the classical complement pathway, as well as an inhibitor of the activated enzymes C1r and C1s. The lack of this protein in serum either as an acquired condition or as a genetic deficiency usually results in chronic or acute angioedema. The goal of this research project is to investigate the physiologic fate of the complexes formed when C1 is activated either in the presence or absence of C1 inhibitor. Our initial efforts have focused on developing sensitive and if possible simpler, more precise hemolytic assays for both C1 inhibitor and proenzyme C1. By taking into account the binding affinities of C1 inhibitor for activated C1r and C1s, a more sensitive assay for C1 inhibitor was devised. Similarly, by modifying the conventional C1 hemolytic titer, a simple yet sensitive and quantitative assay was developed to differentiate unactivated (proenzyme) C1 from activated C1. In exploring the kinetics of the C1-C1 inhibitor interaction we observed that the rate of inactivation of activated C1r2s2 is dependent on the concentration of C1 inhibitor. In addition, our data demonstrate that cell bound C1 is less susceptible to inhibition by C1 inhibitor than is fluid phase activated C1. These findings suggest that those parameters of a substance that limit access of C1 inhibitor to the C1r and C1s enzymes may contribute to the definition of the substance as a C1 activator and may help to explain the seemingly unpredictable clinical manifestation of C1 inhibitor deficiencies.